Sequence-specific recognition is an essential criterion to target and control DNA functions for gene-expression, bio-imaging, diagnostics, therapeutics and biotechnological applications. Over the years, many probes have been developed to target DNA, but there is still a pressing need for developing efficient new probes and therapeutic agents against gene-related diseases. It is a daunting task indeed to design site-specific DNA binding molecules with high affinity and selectivity. To achieve this goal, DNA binding probes ranging from small molecules to large peptides of natural and synthetic origin have been developed. These probes interact with DNA mainly through two binding modes, intercalation and groove binding. Typically, small molecules binding to DNA through intercalation possess the site-specificity of three base pairs that can differentiate only one out of 32 random sequences. Factually, the human genome contains three billion base pairs and the small molecular probe is posed with the astonishingly large number of one billion unique binding sites. To improve the binding specificity of small molecular probes over longer DNA sequences, researchers have shifted their attention towards groove-binding agents.
However, there are several disadvantages/limitations associated with the currently known small molecules/probes which interact with DNA via intercalation and/or groove binding. Some of them are, difficult synthesis processes, non-fluorescent nature limiting their potential applications in biological systems. Further, in the category of fluorescent probes, the blue fluorescence DNA staining probes (DAPI and Hoechst) require excitation in the ultraviolet (UV) region and the prolonged UV illumination is certain to damage cellular DNA. Additionally, propidium iodide and related dyes are intercalators suffering from poor cell-permeability and high toxicity, apart from inducing structural alterations in the target DNA structure, which prohibits their use in biological applications.
The availability of myriad genome data warrants the need for developing efficient and highly predictive molecular tools to probe its organization and functional aspects. Overall, it is clear that sequence-specific targeting of DNA is crucial for studying sequence variation, structural organization and function in the cell nucleus. However, existing probes suffer from various limitations and there is a persistent need to develop probes with superior properties. Especially, fluorescence DNA probes must satisfy the following properties: i) excitation and emission in the longer wavelength region, ii) switch-on fluorescence response, iii) high sequence-specificity, iv) good quantum yield, v) non-toxicity to human cells, and vi) live-cell permeability.
The present disclosure addresses the need by providing potent small molecule probes having superior properties, the process for synthesis of said small molecules and related methods and applications thereof.